Figure 4: Inhibition of OS malignancy by TMIGD3 i1 and A3AR but not i3. | Nature Communications

Figure 4: Inhibition of OS malignancy by TMIGD3 i1 and A3AR but not i3.

From: Genome-wide RNAi screening identifies TMIGD3 isoform1 as a suppressor of NF-κB and osteosarcoma progression

Figure 4

(a) Proliferation assays using SJSA-1 (left) and KHOS/NP (right) cells overexpressing vector control (V), TMIGD3 i1 cDNA (cTi1), TMIGD3 i3 cDNA (cTi3) or A3AR cDNA (cA3). Representative immunoblots showing overexpression of each protein next to the graphs. Error bars: means±s.d. (n=3 independent experiments).*P<0.05 and **P<0.01; two-way analysis of variance (ANOVA). (b) Sphere-formation assays using SJSA-1 and KHOS/NP cells overexpressing V, cTi1, cTi3 or cA3. Graph showing % of sphere formation. Error bars: means±s.d. (n=3 independent experiments). *P<0.05 and **P<0.01, NS, not significant; Student’s t-test (two-tailed). (c) Sphere-formation assays using SJSA-1 cells with or without knockdown of TMIGD3 (T6U, T3) along with overexpression of TMIGD3 i1 (cTi1) or i3 (cTi3). Representative immunoblots below the graphs. Error bars: means±s.d. (n=3 independent experiments). **P<0.01; NS, not significant; Student’s t-test (two-tailed). (d) Sphere-formation assays using SJSA-1 cells with overexpression of A3AR (cA3), knockdown of A3AR (A2a) or knockdown of A3AR (A2a) along with overexpression of A3AR (cA3). Representative immunoblots are below the graphs. Error bars: means±s.d. (n=3 independent experiments). *P<0.05 and **P<0.01; Student’s t-test (two-tailed). (e) Transwell migration assays for 10 h using SJSA-1 (left) and KHOS/NP (right) cells overexpressing V, cTi1, cTi3 or cA3. Graphs showing relative migration to the control (top) and representative images (below). Error bars: means±s.d. (n=3 independent experiments). *P<0.05 and **P<0.01; NS, not significant; Student’s t-test (two-tailed). Scale bar, 25 μm. (f) Tumour growth assays in mice injected with SJSA-1 cells overexpressing V, cTi1, cTi3 or cA3. Cells (1 × 106) were subcutaneously injected into nude mice, and tumour sizes were monitored twice a week for 18 days. Error bars: means±s.d. (n=5 mice per group). Representative images of tumours are next to the graph. **P<0.01; NS, not significant; two-way ANOVA.

Back to article page